Neurons communicate by releasing neurotransmitter, which diffuses to a neighboring cell causing either an excitatory or inhibitory effect by interacting with a receptor. To terminate the signal, released neurotransmitter is deactivated by reuptake into cells or chemical degradation. Thus, the changes in chemical concentrations of neurotransmitters within extracellular space are an indication of the messages sent between neurons. Measurement of these concentration dynamics is an important means of understanding neurotransmission and ultimately the brain and brain diseases. Present methods for monitoring neurotransmitters in vivo include sensors and sampling, fraction collection, and assay of fractions. While sensors are powerful, they are usually limited to measuring one compound at a time and sensors for just a few of the over 100 neurotransmitters have been developed. Sampling approaches are limited by temporal resolution of a few minutes, whereas neurotransmitter concentrations change on the second time scale. In this proposal, we will couple sampling probes with high sensitivity assays, which will allow much more frequent sampling and improve temporal resolution to a few seconds. Specifically, we will: 1) couple sampling on-line with high-speed capillary electrophoresis separations for simultaneous monitoring of up to 6 neurotransmitters or their metabolites, 2) couple sampling off-line with capillary liquid chromatography (LC) for simultaneous monitoring of 11 different amine neurotransmitters, and 3) develop high sensitivity assays (capillary LC or immunoassay) for peptides, which will allow a variety of neuropeptides to be monitored with high temporal resolution. In addition, we will explore a method for discovering new neurotransmitters. Finally, we will use the techniques to explore the regulation of glutamate and aspartate, the principal excitatory neurotransmitters. A greater understanding of glutamate and aspartate regulation is of profound importance since improper regulation is implicated in Parkinson's Disease, epilepsy, and damage associated with stroke.